This project will study the interactions of DNA-binding drugs with DNA. It will investigate the influence of a specific kind of DNA base modification on the stability of drug-DNA complexes. The base modification to be used is halodeoxyuridine substitution -- in which chlorodeoxyuridine, bromodeoxyuridine or iododeoxyuridine are incorporated into DNA in place of thymidine. DNA modified in this way, is normal in every respect except that some (or most) of its thymine methyl groups are replaced by halogen atoms. These halogen atoms, like the methyl groups which they replace, are situated exclusively in the major groove of DNA. Recently, it has been shown that several DNA-binding drugs have an enhanced affinity for halodeoxyuridine-substituted DNAs compared to unsubstituted DNAs. This project will attempt to define which kinds of DNA-binding drugs bind more tightly to halodeoxyuridine-substituted DNAs and why. A large series of different DNA-binding drugs will be surveyed principally by equilibrium dialysis and/or by titration of fluorescence. For each drug, a comparison will be made between the hinding isotherms to unsubstituted DNA and to DNA highly substituted with bromodeoxyuridine. In particular, the binding parameters on each DNA will be evaluated with respect to the number of drug binding sites, the possible presence of more than one class of sites, the affinity constant of the sites and the possible interactions among sites (cooperativity). The drugs to be studied include several anti-tumor and anti-trypanosomal agents as well as some fluorescent drugs currently used in molecular cytogenetics (flow cytometry, visualization of sister chromatid exchanges, chromosome banding procedures, etc.). In studying the interaction of fluorescent drugs with halodeoxyuridine-substituted DNA, a careful distinction will be made between increased or decreased drug binding and quenching or enhancement of fluorescence when the drug is bound to halodeoxyuridine-substituted DNA. As a first approach to that mechanism(s) underlying altered drug binding to halo-substituted DNA, this project will determine whether drugs which cannot make any direct contact with the halogen atoms exhibit altered binding to halo-substituted DNAs.